Comparative Tissue Proteomics of Microdissected Specimens Reveals Novel Candidate Biomarkers of Bladder Cancer

Significance Statement

This manuscript is focused on the biomarker discovery from clinical tissue specimens from bladder cancer patients as well as biomarker verification using several strategies. The results were integrated with a series of papers published by this group of urine proteomes [1-3] to select potential biomarker for further verification studies. Transgelin-2 (TAGLN2) and stathmin 1 (STMN1) proteins were successful verified with higher concentrations in bladder cancer tissue and urine specimens. The integrated results suggest that up-regulated TAGLN2 in bladder tumor cells may be secreted into the urine through microparticles, which in turn causes the higher concentration of urinary TAGLN2.

References:

Chen, Y.T., et al., Discovery of novel bladder cancer biomarkers by comparative urine proteomics using iTRAQ technology. J. Proteome Res., 2010. 9(11): p. 5803-15.
Chen, C.L., et al., Comparative and targeted proteomic analyses of urinary microparticles from bladder cancer and hernia patients. J. Proteome Res., 2012. 11(12): p. 5611-29.
Chen, C.L., et al., Identification of potential bladder cancer markers in urine by abundant-protein depletion coupled with quantitative proteomics. J. Proteomics, 2013. 85C: p. 28-43.

About the author

Dr. Yi-Ting Chen received her Ph.D. degree in Department of Chemistry from the National Tsing Hua University, Taiwan. Before graduation, she went to University of Alberta, Canada as a visiting student in 2001. Following the graduation in 2001, she worked as a research scientist in the Industrial Technology Research Institute, Taiwan to establish a new mass spectrometry lab for numerous proteomic projects. From September 2007, she worked at Molecular Medicine Research Center of Chang Gung University (CGU) as an associated research scientist, with focusing on urinary protein biomarker discovery in bladder cancer. Being an assistant professor in Department of Biomedical Sciences of CGU in 2013, Dr. Chen’s research interests focus on proteomics, metabolomics and systems biology of urological diseases and advanced method development for detection of biomolecules, primarily based on mass spectrometry.

Journal Reference

Mol Cell Proteomics. 2015;14(9):2466-78.

Chien-Lun Chen¹, Ting Chung², Chih-Ching Wu³, Kwai-Fong Ng⁴, Jau-Song Yu⁵, Cheng-Han Tsai⁶, Yu-Sun Chang⁵, Ying Liang², Ke-Hung Tsui¹, Yi-Ting Chen⁷

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From the ‡Department of Urology, Chang Gung Memorial Hospital, Taoyuan, Taiwan; §School of Medicine, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan;
Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; ‖Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
Department of Pathology, Chang Gung Memorial Hospital, Taoyuan, Taiwan;
Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Department of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan. [email protected].

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Abstract

More than 380,000 new cases of bladder cancer are diagnosed worldwide, accounting for ∼150,200 deaths each year. To discover potential biomarkers of bladder cancer, we employed a strategy combining laser microdissection, isobaric tags for relative and absolute quantitation labeling, and liquid chromatography-tandem MS (LC-MS/MS) analysis to profile proteomic changes in fresh-frozen bladder tumor specimens. Cellular proteins from four pairs of surgically resected primary bladder cancer tumor and adjacent nontumorous tissue were extracted for use in two batches of isobaric tags for relative and absolute quantitation experiments, which identified a total of 3220 proteins. A DAVID (database for annotation, visualization and integrated discovery) analysis of dysregulated proteins revealed that the three top-ranking biological processes were extracellular matrix organization, extracellular structure organization, and oxidation-reduction. Biological processes including response to organic substances, response to metal ions, and response to inorganic substances were highlighted by up-expressed proteins in bladder cancer. Seven differentially expressed proteins were selected as potential bladder cancer biomarkers for further verification. Immunohistochemical analyses showed significantly elevated levels of three proteins-SLC3A2, STMN1, and TAGLN2-in tumor cells compared with noncancerous bladder epithelial cells, and suggested that TAGLN2 could be a useful tumor tissue marker for diagnosis (AUC = 0.999) and evaluating lymph node metastasis in bladder cancer patients. ELISA results revealed significantly increased urinary levels of both STMN1 and TAGLN2 in bladder cancer subgroups compared with control groups. In comparisons with age-matched hernia urine specimens, urinary TAGLN2 in bladder cancer samples showed the largest fold change (7.13-fold), with an area-under-the-curve value of 0.70 (p < 0.001, n = 205). Overall, TAGLN2 showed the most significant overexpression in individual bladder cancer tissues and urine specimens, and thus represents a potential biomarker for noninvasive screening for bladder cancer. Our findings highlight the value of bladder tissue proteome in providing valuable information for future validation studies of potential biomarkers in urothelial carcinoma.

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